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Zuo Jin Wan reverses P-gp-mediated drug-resistance by inhibiting activation of the PI3K/Akt/NF-κB pathway.

Sui H, Pan SF, Feng Y, Jin BH, Liu X, Zhou LH, Hou FG, Wang WH, Fu XL, Han ZF, Ren JL, Shi XL, Zhu HR, Li Q - BMC Complement Altern Med (2014)

Bottom Line: We found that ZJW significantly enhanced the sensitivity of chemotherapeutic drugs and increased oxaliplatin (L-OHP)-induced cell apoptosis in a time- and dose-dependent manner.The effect of ZJW in reversing drug-resistance and suppressing phosphorylation of Akt (Ser473) and NF-κB were weakened after treatment with a PI3K/Akt activator in HCT116/L-OHP cells.Our study has provided the first direct evidence that ZJW reverses drug-resistance in human colorectal cancer by blocking the PI3K/Akt/NF-κB signaling pathway, and could be considered as a useful drug for cancer therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Oncology, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China. Lzwf@hotmail.com.

ABSTRACT

Background: Zuo-Jin-Wan (ZJW), a traditional Chinese medicine formula, has been identified to be effective against drug resistance in cancer. In the present study, we investigated the effect of ZJW on acquired oxaliplatin-resistant and the PI3K/Akt/NF-κB pathway in vitro.

Methods: We tested the dose-response relationship of ZJW on reversing drug-resistance by CCK-8 assay and flow cytometry analysis in vitro. The protein expression of P-gp, MRP-2, LRP, and ABCB1 mRNA expression level were evaluated by Western blot and quantitative RT-PCR. The activities of PI3K/Akt/NF-κB pathway were also examined with or without ZJW, including Akt, IκB, p65 and their phosphorylation expression.

Results: We found that ZJW significantly enhanced the sensitivity of chemotherapeutic drugs and increased oxaliplatin (L-OHP)-induced cell apoptosis in a time- and dose-dependent manner. Moreover, both ZJW and a PI3K specific inhibitor (LY294002) suppressed phosphorylation of Akt (Ser473) and NF-κB, which is necessary in the activation of the PI3K/Akt/NF-κB pathway. The effect of ZJW in reversing drug-resistance and suppressing phosphorylation of Akt (Ser473) and NF-κB were weakened after treatment with a PI3K/Akt activator in HCT116/L-OHP cells.

Conclusions: Our study has provided the first direct evidence that ZJW reverses drug-resistance in human colorectal cancer by blocking the PI3K/Akt/NF-κB signaling pathway, and could be considered as a useful drug for cancer therapy.

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ZJW inhibits P-gp expression and the effect of the PI3K/Akt pathway. (A) Western blotting assay was carried out to detect the level of P-gp, LRP, and MRP-2 in HCT116/L-OHP cells treated with LY294002 (20 μM, 2 h), ZJW (50 μg/mL, 48 h), and a combination of ZJW (50 μg/mL, 48 h) and IGF-1 (100 ng/mL, 48 h). GAPDH was used to ensure equal loading of proteins in each lane. (B) Blots were photographed and quantitated; the data are from three independent experiments. (C) Real-time PCR was performed to detect ABCB1 mRNA in vivo. Quantification of ABCB1 mRNA was performed by assigning a value of 1 to the control group treatment with ZJW (50 μg/mL). Statistical difference was analyzed by student’s t-test, **P < 0.01 vs. control group; #P < 0.05 vs. ZJW (50 μg/mL) group. This is a representative result of three experiments with similar results.
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Fig3: ZJW inhibits P-gp expression and the effect of the PI3K/Akt pathway. (A) Western blotting assay was carried out to detect the level of P-gp, LRP, and MRP-2 in HCT116/L-OHP cells treated with LY294002 (20 μM, 2 h), ZJW (50 μg/mL, 48 h), and a combination of ZJW (50 μg/mL, 48 h) and IGF-1 (100 ng/mL, 48 h). GAPDH was used to ensure equal loading of proteins in each lane. (B) Blots were photographed and quantitated; the data are from three independent experiments. (C) Real-time PCR was performed to detect ABCB1 mRNA in vivo. Quantification of ABCB1 mRNA was performed by assigning a value of 1 to the control group treatment with ZJW (50 μg/mL). Statistical difference was analyzed by student’s t-test, **P < 0.01 vs. control group; #P < 0.05 vs. ZJW (50 μg/mL) group. This is a representative result of three experiments with similar results.

Mentions: To further investigate the mechanisms underlying the effect of ZJW and its relationship with the PI3K/Akt pathway, three cell membrane-bound ATP binding cassette (ABC) transporters, P-gp, MRP-2, and LRP, were determined by western blot. Previous study revealed that ZJW administration to MDR cells was accompanied with the downregulation of P-gp, but not other proteins [1]. As shown in Figure 3A and B, LY294002 decreased the expression of P-gp significantly. Moreover, P-gp expression was up-regulated when cells were treated with ZJW and IGF-1. Similarly, we observed a down-regulation in the expression of ABCB1 mRNA in a dose-dependent manner when HCT116/L-OHP cells were treated with ZJW (Figure 3C). These findings support the hypothesis that inhibition of P-gp by ZJW is attributable to suppression of the translational process in the activation of the PI3K/Akt pathway.Figure 3


Zuo Jin Wan reverses P-gp-mediated drug-resistance by inhibiting activation of the PI3K/Akt/NF-κB pathway.

Sui H, Pan SF, Feng Y, Jin BH, Liu X, Zhou LH, Hou FG, Wang WH, Fu XL, Han ZF, Ren JL, Shi XL, Zhu HR, Li Q - BMC Complement Altern Med (2014)

ZJW inhibits P-gp expression and the effect of the PI3K/Akt pathway. (A) Western blotting assay was carried out to detect the level of P-gp, LRP, and MRP-2 in HCT116/L-OHP cells treated with LY294002 (20 μM, 2 h), ZJW (50 μg/mL, 48 h), and a combination of ZJW (50 μg/mL, 48 h) and IGF-1 (100 ng/mL, 48 h). GAPDH was used to ensure equal loading of proteins in each lane. (B) Blots were photographed and quantitated; the data are from three independent experiments. (C) Real-time PCR was performed to detect ABCB1 mRNA in vivo. Quantification of ABCB1 mRNA was performed by assigning a value of 1 to the control group treatment with ZJW (50 μg/mL). Statistical difference was analyzed by student’s t-test, **P < 0.01 vs. control group; #P < 0.05 vs. ZJW (50 μg/mL) group. This is a representative result of three experiments with similar results.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4288643&req=5

Fig3: ZJW inhibits P-gp expression and the effect of the PI3K/Akt pathway. (A) Western blotting assay was carried out to detect the level of P-gp, LRP, and MRP-2 in HCT116/L-OHP cells treated with LY294002 (20 μM, 2 h), ZJW (50 μg/mL, 48 h), and a combination of ZJW (50 μg/mL, 48 h) and IGF-1 (100 ng/mL, 48 h). GAPDH was used to ensure equal loading of proteins in each lane. (B) Blots were photographed and quantitated; the data are from three independent experiments. (C) Real-time PCR was performed to detect ABCB1 mRNA in vivo. Quantification of ABCB1 mRNA was performed by assigning a value of 1 to the control group treatment with ZJW (50 μg/mL). Statistical difference was analyzed by student’s t-test, **P < 0.01 vs. control group; #P < 0.05 vs. ZJW (50 μg/mL) group. This is a representative result of three experiments with similar results.
Mentions: To further investigate the mechanisms underlying the effect of ZJW and its relationship with the PI3K/Akt pathway, three cell membrane-bound ATP binding cassette (ABC) transporters, P-gp, MRP-2, and LRP, were determined by western blot. Previous study revealed that ZJW administration to MDR cells was accompanied with the downregulation of P-gp, but not other proteins [1]. As shown in Figure 3A and B, LY294002 decreased the expression of P-gp significantly. Moreover, P-gp expression was up-regulated when cells were treated with ZJW and IGF-1. Similarly, we observed a down-regulation in the expression of ABCB1 mRNA in a dose-dependent manner when HCT116/L-OHP cells were treated with ZJW (Figure 3C). These findings support the hypothesis that inhibition of P-gp by ZJW is attributable to suppression of the translational process in the activation of the PI3K/Akt pathway.Figure 3

Bottom Line: We found that ZJW significantly enhanced the sensitivity of chemotherapeutic drugs and increased oxaliplatin (L-OHP)-induced cell apoptosis in a time- and dose-dependent manner.The effect of ZJW in reversing drug-resistance and suppressing phosphorylation of Akt (Ser473) and NF-κB were weakened after treatment with a PI3K/Akt activator in HCT116/L-OHP cells.Our study has provided the first direct evidence that ZJW reverses drug-resistance in human colorectal cancer by blocking the PI3K/Akt/NF-κB signaling pathway, and could be considered as a useful drug for cancer therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Oncology, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China. Lzwf@hotmail.com.

ABSTRACT

Background: Zuo-Jin-Wan (ZJW), a traditional Chinese medicine formula, has been identified to be effective against drug resistance in cancer. In the present study, we investigated the effect of ZJW on acquired oxaliplatin-resistant and the PI3K/Akt/NF-κB pathway in vitro.

Methods: We tested the dose-response relationship of ZJW on reversing drug-resistance by CCK-8 assay and flow cytometry analysis in vitro. The protein expression of P-gp, MRP-2, LRP, and ABCB1 mRNA expression level were evaluated by Western blot and quantitative RT-PCR. The activities of PI3K/Akt/NF-κB pathway were also examined with or without ZJW, including Akt, IκB, p65 and their phosphorylation expression.

Results: We found that ZJW significantly enhanced the sensitivity of chemotherapeutic drugs and increased oxaliplatin (L-OHP)-induced cell apoptosis in a time- and dose-dependent manner. Moreover, both ZJW and a PI3K specific inhibitor (LY294002) suppressed phosphorylation of Akt (Ser473) and NF-κB, which is necessary in the activation of the PI3K/Akt/NF-κB pathway. The effect of ZJW in reversing drug-resistance and suppressing phosphorylation of Akt (Ser473) and NF-κB were weakened after treatment with a PI3K/Akt activator in HCT116/L-OHP cells.

Conclusions: Our study has provided the first direct evidence that ZJW reverses drug-resistance in human colorectal cancer by blocking the PI3K/Akt/NF-κB signaling pathway, and could be considered as a useful drug for cancer therapy.

Show MeSH
Related in: MedlinePlus